The challenge of membrane protein structure determination remains largely unmet. Thousands of water-soluble protein structures have been solved, while, in contrast, only a handful of membrane protein structures have been solved. Yet estimates suggest that 25-40 percent of the coding regions of genomes from higher organisms code for membrane proteins. As much as 60 percent of the drug targets of the pharmaceutical industry are a family of membrane proteins called G-protein coupled receptors, yet there are no X-ray crystal structures of any of these drug targets. The long-term goal of this project is to construct a path around the current problems in membrane protein structure determination through the development of a new approach to this problem. While this new approach does not replace X-ray crystallography, it does offer structural information where crystallization suitable for diffraction studies has proven impossible. It also offers the possibility of working on proteins whose size would make them unsuitable subjects for standard NMR techniques, particularly where the effective size of the membrane protein is increased by inclusion in a detergent micelle or in a lipid bicelle. In this R21 application, we focus on a difficult challenge: the solution of the structure for a 12 TM membrane protein that has so far proven refractory to X-ray crystallography. We will use what we have learned from the solution of the structures of two 7 TM membrane proteins and determine whether the same approach can be used to solve the three dimensional structure of a much larger membrane protein, the lac permease. This structure determination of the lac permease will prove very helpful in understanding the mechanism of transport for this and related proteins. Success of this project will provide a powerful new tool for proteomics that can relatively rapidly produce useful structural information for many families of membrane proteins.